Perk-dependent repression of miR-106b-25 cluster is required for ER stress-induced apoptosis

Activation of the unfolded protein response sensor PKR-like endoplasmic reticulum kinase (Perk) attenuates endoplasmic reticulum (ER) stress levels. Conversantly, if the damage is too severe and ER function cannot be restored, this signaling branch triggers apoptosis. Bcl-2 homology 3-only family member Bim is essential for ER stress-induced apoptosis. However, the regulatory mechanisms controlling Bim activation under ER stress conditions are not well understood. Here, we show that downregulation of the miR-106b-25 cluster contributes to ER stress-induced apoptosis and the upregulation of Bim. Hypericin-mediated photo-oxidative ER damage induced Perk-dependent cell death and led to a significant decrease in the levels of miRNAs belonging to miR-106b-25 cluster in wild-type (WT) but not in Perk−/− MEFs. Further, we show that expression of miR-106b-25 and Mcm-7 (host gene of miR-106b-25) is co-regulated through the transcription factors Atf4 (activating transcription factor 4) and Nrf2 (nuclear factor-erythroid-2-related factor 2). ER stress increased the activity of WT Bim 3′UTR (untranslated region) construct but not the miR-106b-25 recognition site-mutated Bim 3′UTR construct. Overexpression of miR-106b-25 cluster inhibits ER stress-induced cell death in WT but did not confer any further protection in Bim-knockdown cells. Further, we show downregulation in the levels of miR-106b-25 cluster in the symptomatic SOD1G86R transgenic mice. Our results suggest a molecular mechanism whereby repression of miR-106b-25 cluster has an important role in ER stress-mediated increase in Bim and apoptosis

Steel-based applications in earthquake-prone areas

Steel-Earth project aims at distributing among technicians, engineers, design companies and standardization bodies the results of three past RFCS projects (Steel-Retro [3], Opus [2] and PrecaSteel [1]), providing useful tools for the design and for the retrofit of existing buildings. Technical documents and practical applications to case studies, regarding design of steel and composite steel/concrete buildings and innovative steel-based techniques for the retrofit of existing r.c. and masonry constructions, have been elaborated and collected into a volume distributed during the final workshop of the dissemination project. Pre-normative and background documents concerning the design of steel and composite structures and the rehabilitation of existing constructions have been prepared. A lot of attention has been paid to the analysis of the influence of overstrength factors on the seismic design of steel and composite structures. The prepared documents have been distributed to the attending people and to the members of WG 2 (CEN/TC 250/SC 8/WG 2 “Steel and Composite Structures”) during the final workshop of the project. Technical sheets, working examples and background documents have been translated into several languages (German, French, Italian, Romanian and Greek) and are free available on the website of the project (https://www.steelconstruct.com/site/), where information regarding Steel-Earth are also presented.11 Workshops in Italy, Greece, Germany, Belgium, Portugal, Spain and Romania and 5 conferences in Emilia-Romagna have been organized, as well as 2 practical courses for engineers and academic people in Pavia (Italy). Flash-drives with the technical documents and applications elaborated in Steel-Earth have been distributed to the attending people

Living with a Crucial Decision: A Qualitative Study of Parental Narratives Three Years after the Loss of Their Newborn in the NICU

BACKGROUND: The importance of involving parents in the end-of-life decision-making-process (EOL DMP) for their child in the neonatal intensive care unit (NICU) is recognised by ethical guidelines in numerous countries. However, studies exploring parents' opinions on the type of involvement report conflicting results. This study sought to explore parents' experience of the EOL DMP for their child in the NICU. METHODS: The study used a retrospective longitudinal design with a qualitative analysis of parental experience 3 years after the death of their child in four NICUs in France. 53 face-to-face interviews and 80 telephone interviews were conducted with 164 individuals. Semi-structured interviews were conducted to explore how parents perceived their role in the decision process, what they valued about physicians' attitudes in this situation and whether their long-term emotional well being varied according to their perceived role in the EOL DMP. FINDINGS: Qualitative analysis identified four types of perceived role in the DMP: shared, medical, informed parental decision, and no decision. Shared DM was the most appreciated by parents. Medical DM was experienced as positive only when it was associated with communication. Informed parental DM was associated with feelings of anxiousness and abandonment. The physicians' attitudes that were perceived as helpful in the long term were explicit sharing of responsibility, clear expression of staff preferences, and respectful care and language toward the child. INTERPRETATION: Parents find it valuable to express their opinion in the EOL DMP of their child. Nonetheless, they do need continuous emotional support and an explicit share of the responsibility for the decision. As involvement preferences and associated feelings can vary, parents should be able to decide what role they want to play. However, our study suggests that fully autonomous decisions should be misadvised in these types of tragic choices

The Ubiquitin Peptidase UCHL1 Induces G0/G1 Cell Cycle Arrest and Apoptosis Through Stabilizing p53 and Is Frequently Silenced in Breast Cancer

Background: Breast cancer (BrCa) is a complex disease driven by aberrant gene alterations and environmental factors. Recent studies reveal that abnormal epigenetic gene regulation also plays an important role in its pathogenesis. Ubiquitin carboxyl- terminal esterase L1 (UCHL1) is a tumor suppressor silenced by promoter methylation in multiple cancers, but its role and alterations in breast tumorigenesis remain unclear. Methodology/Principal Findings: We found that UCHL1 was frequently downregulated or silenced in breast cancer cell lines and tumor tissues, but readily expressed in normal breast tissues and mammary epithelial cells. Promoter methylation of UCHL1 was detected in 9 of 10 breast cancer cell lines (90%) and 53 of 66 (80%) primary tumors, but rarely in normal breast tissues, which was statistically correlated with advanced clinical stage and progesterone receptor status. Pharmacologic demethylation reactivated UCHL1 expression along with concomitant promoter demethylation. Ectopic expression of UCHL1 significantly suppressed the colony formation and proliferation of breast tumor cells, through inducing G0/G1 cell cycle arrest and apoptosis. Subcellular localization study showed that UCHL1 increased cytoplasmic abundance of p53. We further found that UCHL1 induced p53 accumulation and reduced MDM2 protein level, and subsequently upregulated the expression of p21, as well as cleavage of caspase3 and PARP, but not in catalytic mutant UCHL1 C90Sexpressed cells

Human Embryonic Stem Cells Express Elevated Levels of Multiple Pro-Apoptotic BCL-2 Family Members

Two of the greatest challenges in regenerative medicine today remain (1) the ability to culture human embryonic stem cells (hESCs) at a scale sufficient to satisfy clinical demand and (2) the ability to eliminate teratoma-forming cells from preparations of cells with clinically desirable phenotypes. Understanding the pathways governing apoptosis in hESCs may provide a means to address these issues. Limiting apoptosis could aid scaling efforts, whereas triggering selective apoptosis in hESCs could eliminate unwanted teratoma-forming cells. We focus here on the BCL-2 family of proteins, which regulate mitochondrial-dependent apoptosis. We used quantitative PCR to compare the steady-state expression profile of all human BCL-2 family members in hESCs with that of human primary cells from various origins and two cancer lines. Our findings indicate that hESCs express elevated levels of the pro-apoptotic BH3-only BCL-2 family members NOXA, BIK, BIM, BMF and PUMA when compared with differentiated cells and cancer cells. However, compensatory expression of pro-survival BCL-2 family members in hESCs was not observed, suggesting a possible explanation for the elevated rates of apoptosis observed in proliferating hESC cultures, as well as a mechanism that could be exploited to limit hESC-derived neoplasms

Tumor Transcriptome Sequencing Reveals Allelic Expression Imbalances Associated with Copy Number Alterations

Due to growing throughput and shrinking cost, massively parallel sequencing is rapidly becoming an attractive alternative to microarrays for the genome-wide study of gene expression and copy number alterations in primary tumors. The sequencing of transcripts (RNA-Seq) should offer several advantages over microarray-based methods, including the ability to detect somatic mutations and accurately measure allele-specific expression. To investigate these advantages we have applied a novel, strand-specific RNA-Seq method to tumors and matched normal tissue from three patients with oral squamous cell carcinomas. Additionally, to better understand the genomic determinants of the gene expression changes observed, we have sequenced the tumor and normal genomes of one of these patients. We demonstrate here that our RNA-Seq method accurately measures allelic imbalance and that measurement on the genome-wide scale yields novel insights into cancer etiology. As expected, the set of genes differentially expressed in the tumors is enriched for cell adhesion and differentiation functions, but, unexpectedly, the set of allelically imbalanced genes is also enriched for these same cancer-related functions. By comparing the transcriptomic perturbations observed in one patient to his underlying normal and tumor genomes, we find that allelic imbalance in the tumor is associated with copy number mutations and that copy number mutations are, in turn, strongly associated with changes in transcript abundance. These results support a model in which allele-specific deletions and duplications drive allele-specific changes in gene expression in the developing tumor

Development and initial validation of the EDIN scale, a new tool for assessing prolonged pain in preterm infants

OBJECTIVE—To develop and validate a scale suitable for use in clinical practice as a tool for assessing prolonged pain in premature infants.
METHODS—Pain indicators identified by observation of preterm infants and selected by a panel of experts were used to develop the EDIN scale (Échelle Douleur Inconfort Nouveau-Né, neonatal pain and discomfort scale). A cohort of preterm infants was studied prospectively to determine construct validity, inter-rater reliability, and internal consistency of the scale.
RESULTS—The EDIN scale uses five behavioural indicators of prolonged pain: facial activity, body movements, quality of sleep, quality of contact with nurses, and consolability. The validation study included 76 preterm infants with a mean gestational age of 31.5weeks. Inter-rater reliability was acceptable, with a κ coefficient range of 0.59-0.74. Internal consistency was high: Cronbach's α coefficients calculated after deleting each item ranged from 0.86 to 0.94. To establish construct validity, EDIN scores in two extreme situations (pain and no pain) were compared, and a significant difference was observed.
CONCLUSIONS—The validation data suggest that the EDIN is appropriate for assessing prolonged pain in preterm infants. Further studies are warranted to obtain further evidence of construct validity by comparing scores in less extreme situations.